Kaposi's sarcoma (KS)-associated herpesvirus (KSHV or HHV8) is the causative agent of KS, primary effusion lymphoma (PEL) and multicentric Castleman's disease. These tumors occur in AIDS and other immunocompromised states. There are no specific therapies for these diseases. KS is the leading AIDS malignancy, and is epidemic in sub Saharan Africa. KS often involves the oral cavity and can disseminate to visceral organs. KSHV latently infects tumor cells and viral genomes persist as circular, multiple copy, extrachromosomal, episomes (plasmids). In order to persist in proliferating cells, episomes must first replicate and then efficiently segregate to daughter nuclei. Tumor cell viability is dependent on latent KSHV infection. The latency-associated nuclear antigen (LANA) mediates KSHV episome persistence and this function is essential for virus survival. Therefore, LANA function is of fundamental importance. LANA represents the Achilles heel of KSHV as its disruption is lethal to the virus. LANA is necessary and sufficient for KSHV episome maintenance in the absence of other virus genes. Episome persistence is comprised of two components: 1) replication of viral DNA and 2) segregation of the replicated episomes to progeny cell nuclei. LANA mediates both these functions. C-terminal LANA binds specific sequence in KSHV terminal repeat (TR) DNA to mediate DNA replication. LANA tethers TR DNA to mitotic chromosomes to segregate genomes to daughter nuclei. To form a molecular tether, the C-LANA DNA binding domain (DBD) binds TR DNA and N-LANA simultaneously binds histones H2A/H2B at the folded portion of the nucleosome. In addition to N- and C-LANA, internal LANA sequence is also critical for episome persistence. We have discovered two distinct internal LANA regions, one comprised of unique sequence, and one within repetitive sequence, that are critical for LANA mediated episome persistence. The unique sequence contains three distinct functional motifs, whereas the repetitive region is homologous to a host cell ?reader? sequence. Both of these regions will be investigated in this work. This work will use a detailed, in depth approach to investigate each of the recently identified LANA functional sequences. Experiments will use a rigorously developed panel of assays to identify the functional motif and determine the mechanism through which the unique sequence mediates episome persistence. The repetitive LANA effector sequence homologous to host ?reader? sequence will be investigated to determine the mechanism through which it exerts its function. This work will provide important insight into LANA?s fundamental function of episome maintenance, and may lead to novel strategies to prevent or treat KSHV associated malignancies.